PROJECT SUMMARY Neurofibromatosis type 2 (NF2) is a tumor disorder characterized by development of bilateral vestibular schwannomas also called acoustic neuromas. 98% of all NF2 patients experience partial to complete loss of hearing. Treatment for NF2 is balanced between monitoring tumor growth and the slow but progressive loss of hearing with surgical removal of larger tumors impinging on brainstem function and complete and permanent deafness. Cochlear and auditory brainstem implants have been used to partially restore hearing in a subset of patients with varying success. Therapeutics that slow or reverse tumor growth whilst maintaining hearing are currently lacking. This proposal tests the hypothesis that correcting the cytoskeletal defects in supernumerary schwannoma cells lacking function of the nf2 gene product, schwannomin/merlin, will allow these cells to interact with axons and receive cues promoting their differentiation and/or apoptosis. Identifying proteins that directly bind actin and regulate Schwann cell morphology is of the utmost importance. These proteins can serve as targets for drugs that will repair actin dynamics in schwannoma cells and restore axonal contact. Alternatively, drugs that modify actin regulatory proteins could promote cell death as a result of failed cytokinesis and mitotic spindle organization. One function of schwanomin/merlin is to inhibit Cdc42/Rac activation of p21 activated kinase (PAK). We will investigate LIM kinase (LIMK) and cofilin, terminal targets in a PAK signaling pathway. LIMK is a substrate for PAK, thus its activity is predicted to be high in schwannomas. Cofilin is a ubiquitously expressed actin-binding factor that depolymerizes f-actin and creates nucleation sites for new actin polymerization. Cofilin's function is inhibited by phosphorylation on serine-3 by LIMK. Our preliminary studies demonstrate that LIMK and cofilin modulate actin dynamics and function in Schwann cells. Moreover, our results suggest that LIMK and cofilin act down-stream of Schwannomin/merlin. We propose studies to: 1) identify the role of these proteins in controlling actin polymerization and cellular function in normal rat Schwann cells, 2) establish an in vitro model for NF2 using nf2ex2deleted mouse SCs to determine if inactivation of schwannomin/merlin leads to de-regulation of LIMK and cofilin activity and loss of SC function, and 3) determine if modulators of LIMK and cofilin restore the morphology and function of nf2ex2deleted SCs. These studies are initial steps in validating LIMK and/or cofilin as drug targets for development of an effective treatment for NF2 aimed at preserving hearing.